International Journal of Pharmaceutics 597 (2021) 120312

Contents lists available at ScienceDirect

International Journal of Pharmaceutics

journal homepage: www.elsevier.com/locate/ijpharm

Effect of co-milling on dissolution rate of poorly soluble drugs

Michaela Slámová a, *, Kateřina Prausová a, Julie Epikaridisová a, Jana Brokešová b,
Martin Kuentz c, Jan Patera a, Petr Zámostný a
a
Department of Organic Technology, UCT Prague, Technická 5, 166 28 Prague 6, Dejvice, Czech Republic
b
Faculty of Pharmacy, Department of Pharmaceutical Technology, Charles University, Ak. Heyrovského 1203, 500 05 Hradec Králové, Czech Republic
c
University of Applied Sciences and Arts Northwestern Switzerland, Institute of Pharmaceutical Technology, Gründenstr. 40, CH-4132 Muttenz, Switzerland

A R T I C L E I N F O A B S T R A C T

Keywords: Co-milling of a drug with a co-former is an efficient technique to improve the solubility of drugs. Besides the
Co-milling particle size reduction, the co-milling process induces a structural disorder and the creation of amorphous re­
Glass forming ability gions. The extent of drug solubility enhancement is dependent on the proper choice of co-milling co-former. The
Indomethacin
aim of this work was to compare the effects of different co-formers (meglumine and polyvinylpyrrolidone) on the
Mefenamic acid
dissolution rates of glass forming (indomethacin) and non-glass forming (mefenamic acid) model drugs. A
Dissolution rate
positive impact of the co-milling on the dissolution behavior was observed in all co-milled mixtures, even if no
substantial amorphization was observed. While meglumine exhibited pronounced effects on the dissolution rate
of both drugs, the slightest enhancement was observed in mixtures with polyvinylpyrrolidone. The evaluation of
specific release rate revealed the surface activation of drug particle is responsible for improving the dissolution
rate of both drug types, but for the glass former, this surface activation could be persistent while maintaining a
high dissolution rate even until a high fraction of drug is released. Our results, therefore, indicate that adequate
co-former choice and consideration of drug glass forming ability are important for a successful co-milling
approach to poorly water-soluble drugs.

1. Introduction Particle size reduction is commonly used because smaller particles

Bioavailability is an essential property of any drug and plays a sig­
nificant role in its successful per os administration, which remains the
most common administration route. However, many newly developed
drugs have very low to almost no solubility in aqueous media. Hence,
improving the solubility and bioavailability of new formulations is one
of the major challenges facing the pharmaceutical industry (Kawabata
et al., 2011).
Several strategies have been applied to the improvement of drug
solubility, including cyclodextrin complex formation, nanocrystal for­
mation, prodrug development, particle size reduction, or drug
amorphization (Borba et al., 2016; Li et al., 2017; Maggi et al., 2015).
Cyclodextrin complex or nanocrystals can be prepared in the laboratory,
however, scale-up in the industry poses technical challenges (Kawabata
et al., 2011).
have a larger surface area, enabling the drug to be dissolved more easily
(Caron et al., 2011; Isaac et al., 2016). This method involves the use of a
drug milling technology (ball mill, hammer mill, micronizer, or nano
mill), and especially promising is co-milling of a drug with a feasible co-
former (Mura et al., 2002; Szafraniec et al., 2017). Co-milling is an
energy-intensive process capable of producing well mixed drug-
excipient blends with a possible alteration of the degree of crystal­
linity (Bahl et al., 2008; Maggi et al., 2015, 2013). The interaction be­
tween the drug and the hydrophilic excipient increases wettability,
thereby enhancing the dissolution and possibly also bioavailability of a
drug (Loh et al., 2014).
Drug amorphization is another key approach to improve apparent
solubility and dissolution rates. Amorphous solid dispersions can be
produced by a variety of manufacturing techniques, including spray
drying, freeze drying, melt extrusion, and again co-milling (Kawabata

Abbreviations: COM, co-milled mixture; FTIR, Fourier transform infrared spectroscopy; HPLC, high-pressure liquid chromatography; IND, indomethacin; MA,
mefenamic acid; MG, meglumine; nGF, non-glass formers; PM, physical mixture; PVP, polyvinylpyrrolidone; SEM, scanning electron microscopy; sGF, stable glass
former; uGF, unstable glass former; USP4, flow-through cell apparatus; XRD, powder X-ray diffraction.
* Corresponding author.
E-mail address: slamovam@vscht.cz (M. Slámová).

https://doi.org/10.1016/j.ijpharm.2021.120312
Received 13 October 2020; Received in revised form 20 January 2021; Accepted 21 January 2021
Available online 1 February 2021
0378-5173/© 2021 Elsevier B.V. All rights reserved.
M. Slámová et al.
et al., 2011; Löbmann et al., 2014; Varghese and Ghoroi, 2017). How­
ever, an amorphous form (also known as a glass form) has higher energy
than its corresponding crystalline form and is thermodynamically less
stable, meaning that is often reverts to a more stable crystalline form
(Caron et al., 2011; Lim et al., 2013b; Löbmann et al., 2014). Baird et al.
(Baird et al., 2010), Wyttenbach et al. (Wyttenbach et al., 2016) and
Alhalaweh et al. (Alhalaweh et al., 2014) helpfully divided drugs into
three groups based on their glass forming ability: stable glass formers
(sGFs) transform easily into an amorphous solid and have the least
tendency to recrystallize; non-glass formers (nGF) show barely a vitri­
fication and recrystallize upon temperature reduction; unstable (uGFs)
can be converted into an amorphous form, but they recrystallize quickly
above their glass transition temperature.
Many studies have investigated the use of co-milled mixtures of one
the glass forming types of drugs and a specific excipient. These studies
have focused on poorly soluble drugs, such as indomethacin (Bahl et al.,
2008; Patterson et al., 2007), piroxicam (Koh et al., 1986), naproxen
(Gupta et al., 2003), meloxicam (Kürti et al., 2011), nimesulide, pro­
bucol (Li et al., 2017), furosemide (Kaminska et al., 2013; Nkansah et al.,
2013) amongst many others (Varghese and Ghoroi, 2017). In general,
hydrophilic or amphiphilic polymers and poorly water-soluble non­
polymers have been used for milling with sGFs, and polymeric excipients
with nGFs; uGFs have been co-milled with an entire range of excipients.
However, while these researchers often tested multiple excipients, they
did so mostly without addressing the ability to form a glass and/or by
using only a drug of a single glass forming type. Accordingly, no direct
comparison has been made of excipient use with model sGF and nGF
drugs.
Therefore, we aim to study co-milling with various excipients to
improve the dissolution rates of model poorly soluble drugs with either
stable (sGFs) or poor (nGF) glass forming ability. Mefenamic acid was
chosen as a typical nGF, while indomethacin was a model drug of an sGF
that more easily converts to an amorphous form with comparatively
good kinetic stability. The drugs were co-milled with different co-
formers to form binary mixtures having various drug to co-former ra­
tios. Co-formers were selected based on acid-base properties. Meglumine
was used as a basic additive with high solubility, and poly­
vinylpyrrolidone (PVP) was used as a neutral co-former (hydrophilic
model polymer). Binary mixtures were milled in the planetary ball mill,
and dissolution tests were conducted in the flow-through cell apparatus
(USP4). After that, we compared these co-formers. Interestingly, all the
co-milled mixtures showed improved dissolution behavior, suggesting
that the method applied might be suitable for both poorly water-soluble
sGFs and nGFs.
2. Material and methods
2.1. Materials
Indomethacin (IND) and Mefenamic acid (MA) were obtained from
Sigma-Aldrich (Prague, Czech Republic). Povidone 25 (PVP) and
meglumine (MG) were purchased from Zentiva, k.s. (Prague, Czech
Republic). Methanol (LC/MS grade) and Acetonitrile (LC/MS grade)
were obtained from Fisher Chemical (Prague, Czech Republic). Acetic
acid p.a., Sodium hydroxide and Potassium dihydrogen phosphate p.a.
were purchased from Penta (Prague, Czech Republic).
2.2. Preparation of PM and COM
Firstly, physical mixtures (PMs), as reference material, were pre­
pared by mixing IND or MA with selected co-formers in 1:1, 1:2 and 1:4
drug to carrier ratio (w/w) using 3D blender Turbula TF2 (W.A. Bach­
ofen, Basel, Switzerland) at 50 rpm and 15 min. Subsequently, the
mixtures were mechanically processed in a Planetary ball mill Retsch
PM 100 CM (Haan, Germany) using 5 stainless steel balls (diameter 10
mm) for 20 min at 250 rpm to create co-milled mixtures (COM).
2
International Journal of Pharmaceutics 597 (2021) 120312
2.3. Flow-through cell dissolution tests
Dissolution was tested in the flow-through cell apparatus (USP4)
using an open-loop system Sotax Dissotest CE1 (Sotax, Basel,
Switzerland) and a piston pump Sotax CY 1 (Sotax, Basel, Switzerland)
at 37 ◦ C. The cell with a 12 mm diameter and a 32 mm height was used
for approximately 20 mg of powder. The cell was tempered in a water
bath at 37 ◦ C. The phosphate buffer (pH 6.8) was used as a dissolution
medium, which was pumped through the system with the volumetric
flow of 16 ml⋅min− 1. The samples were collected at short time intervals:
0.5; 1; 1.5; 2; 2.5; 3; 4; 5; 6; 7; 8; 9; 10; 15; 20; 25; 30; 35; 40; 45 min and
analysed using high-performance liquid chromatography, HPLC. The
duration of the experiments was 45 min in the case of mefenamic acid
and 30 min in the case of indomethacin. The duration of the experiment
was different for both substances, because in the case of indomethacin
no detectable concentration of IND was observed in the samples after 30
min and it was therefore not necessary to continue the experiment.
The mass flow ṁ (mg⋅min− 1) of the dissolved drug from the disso­
lution cell is the product of mass concentration c (mg⋅ml− 1) and volu­
metric flow F (ml⋅min− 1) of the dissolution media (Eq. (1)):
ṁ = c∙F (1)
Assuming the small volume of the dissolution cell and plug flow of
the liquid via the outlet tubing, the mass flow from the dissolution cell
approximates the mass flow through the solid–liquid phase interface. A
relative release rate of drug (min− 1) is defined as the ratio of mass flow
and initial mass of drug in a mixture (Eq. (2)):
ṁ c∙F
= (2)
m0 m0
It provides a comparable quantity to express the release rate inde­
pendently of the total initial load of the drug in the dissolution cell.
A released fraction of drug w (-) is calculated using the trapezoid
integration rule as follows (Eq. (3)):
mi− 1 +mi
wi = wi− 1 + (ti − ti− 1 )∙
m0
(3)
2
where the ti represents the time of i-th sample collection.
A ratio of mass flow and residual mass of drug is expressed (Eq.4) by
specific release rate μ’ (min− 1):
′
μ =
ṁ
=
ṁ
(4)
mres m0 ∙(1 − w)
√̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅
which is modified by the scale factor of (1 − w)2 instead of (1 – w) to get
3
the surface-equivalent specific release rate μ (min− 1) according to the Eq. (5):
ṁ
(5)
m0
μ = √̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅
̅
(1 − w)2
3
√̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅̅
where (1 − w)2 ∙m0 represents the surface equivalent residual mass of
3
the non-dissolved drug.
2.4. HPLC analysis
The concentration of IND and MA was measured using HPLC Dionex
Ultimate 3 000 (Sunnyvale, California, USA) with a C18 Kinetex® Col­
umn 5 μm 100 Å (Phenomenex® Torrance, CA, USA). Based on the
absorption spectra of both drugs, the wavelengths were chosen: 285 nm
for mefenamic acid, and 267 nm for indomethacin. For indomethacin,
the mobile phase consisted of acetonitrile/water/acetic acid 90/60/5
(v/v), while for mefenamic acid, acetonitrile/0.1 M acetic acid 70/30
(v/v) was used. The flow rate was 1 ml⋅min− 1, the temperature of the
oven 30 ◦ C and the injected volume of the sample 10 μl.
M. Slámová et al. International Journal of Pharmaceutics 597 (2021) 120312

2.5. Powder X-ray diffraction (XRD)
The XRD data were collected at room temperature on a Bruker D2
Phaser powder diffractometer with parafocusing Bragg-Brentano ge­
ometry (Bruker, Billerica, MA, USA) using CuKα radiation (k = 0,15418,
U = 30 kV, I = 10 mA), and LYNXEYE_XE 1D detector. Samples were
scanned in steps 0.02◦ from 5◦ to 80◦ (diffraction angle 2θ). Data eval­
uation was performed in the software package HighScorePlus.
2.6. Scanning electron microscopy (SEM)
All samples were taken with FE MIRA II LMU scanning electron
microscope (Tescan, Brno, Czech Republic). First, the powder was fixed
to a graphite double-sided adhesive tape and then covered with 5 nm
gold combined sprayer Quorum Q150R ES (Quorum Technologies Ltd.,
Laughton, UK). The images were performed at magnifications 1 000,
2 000 and 3 000.
2.7. Optical microscope
All samples were taken with a Nikon Research Stereo Microscope
SMZ 18 (Nikon Corporation, Tokyo, Japan). NIS-Elements AR software
was used for image analysis. The images were performed at magnifica­
tions of 30 times.
2.8. Static laser light scattering
Particle size distribution was measured using Malvern Mastersizer
3000 with Aero D dry unit (Malvern Instruments Ltd., Worcestershire,
United Kingdom). 4mW He-Ne 632.8 nm red light source and 10 mW
LED 470 nm blue light source was used. The range of measurement was
set from 0.1 to 1 000 µm. Small spoon of dry sample was placed into the
cell and particle sizes of x10 (µm), x50 (µm) and x90 (µm) were detected.
Span value characerizing the width of particle size distribution was
evaluated as well. Mie theory of static light scattering was utilized
during the measurement. All samples were measured 5 times (5 scans)
and the distribution was created as an average value.
2.9. Fourier transform infrared (FTIR) spectroscopy
A Nicolet iS50 FTIR (Thermo Scientific Waltham, MA, USA) was used
to investigate potential interactions between drugs and excipients.
Measurements were carried out in attenuated total reflection (ATR)
mode, in the range of 4000–400 cm− 1 with a resolution of 2 cm− 1.
3. Results and discussion
3.1. Flow-through cell dissolution
The flow-through cell apparatus for powder was used to study the
dissolution properties of IND and MA from PMs and COMs. Results of the
measurements with MG are depicted in Fig. 1 and show the percentage
of the released drug over time. In the case of PM IND-MG, it was evident
that as the content of MG increased, also the released amount of IND
increased. After co-milling, the dissolution rates were much higher than
for pure or milled IND (Table 1) and much higher than for corresponding
PMs. It was also observed that the released amount of IND was inde­
pendent of the MG content, meaning that even the smallest 1:1 (w/w)
MG addition ratio had a similar effect to the highest 1:4 (w/w) one.
In the case of MA, the milling did not produce any significant impact
on the dissolution rate. Clearly, the system was lacking the propensity to
create disorder upon mechanical treatment (as a non-glass forming
compound) and effective particle size reduction did not occurred. These
observations were confirmed with XRD where almost no change was
observed after the mechanical processing (see below). There was no
difference in dissolution patterns between the pure and milled MA.

A) B)
120 120

100 100

80 80
wIND (%)

wIND (%)

60 60

40 1:4
40 1:4
1:2 1:2
20 1:1 20 1:1
IND milled IND milled
IND pure IND pure
0 0
0 5 10 15 20 25 30 0 5 10 15 20 25 30

C) t (min) t (min)
D)
100 1:4
100
1:2
1:1
80 MA milled
80
MA pure
wMA (%)
wMA (%)

60 60
1:4
1:2
40 40 1:1
MA milled
MA pure
20 20

0 0
0 5 10 15 20 25 30 35 40 45 0 5 10 15 20 25 30 35 40 45

t (min) t (min)

Fig. 1. Dissolution profiles of mixtures of indomethacin (IND) or mefenamic acid (MA) with meglumine (MG) prepared by physical mixing (PM) or co-milling
(COM): IND/MG PMs (A), IND/MG COMs (B), MA/MG PMs (C) and MA/MG COMs (D).

3
M. Slámová et al. International Journal of Pharmaceutics 597 (2021) 120312

Table 1
Maximal values of relative release rates of drug.
Co-former Drug: Co-former ratio (w/w) Mixture type rMAX (min¡1)

Indomethacin Mefenamic acid

Pure drug Pure drug 0.066 ± 0.011 0.003 ± 0.001

Milled drug 0.185 ± 0.023 0.004 ± 0.001
Meglumine 1:1 PM 0.810 ± 0.398 0.169 ± 0.060
COM 1.225 ± 0.438 0.243 ± 0.075
1:2 PM 1.608 ± 0.310 0.536 ± 0.037
COM 1.676 ± 0.290 0.326 ± 0.008
1:4 PM 1.819 ± 0.441 0.928 ± 0.001
COM 1.849 ± 0.527 1.183 ± 0.103
Polyvinylpyrrolidone 1:1 PM 0.049 ± 0.010 0.008 ± 0.003
COM 0.211 ± 0.019 0.005 ± 0.001
1:2 PM 0.105 ± 0.002 0.023 ± 0.014
COM 0.229 ± 0.024 0.012 ± 0.003
1:4 PM 0.037 ± 0.018 0.051 ± 0.018
COM 0.321 ± 0.072 0.032 ± 0.003

These results implied that reducing the particle size by milling may not
be the most promising approach to increase the MA dissolution rate.
The co-former used for co-milling with the drug was expected to play
an important role. Such importance has already been confirmed in the
literature for other drugs, e.g. probucol and bicalutamide (Li et al., 2017;
Szafraniec et al., 2017), but it was not systematically discussed in rela­
tion to the drug glass forming ability. Indeed, the presence of MG in the
mixtures enhanced the dissolution behavior of MA enormously in
comparison to the MA milled alone. A MG fraction in the binary mixture
played an important role in improving the dissolution behavior. In the
case of PM MA-MG, the results showed that with higher ratios of MG in
the mixture, also higher amounts of dissolved MA were achieved. MA
released most rapidly from the 1:4 (w/w) mixture. Thus, an interactive
mixture can be anticipated from which the drug substance was released
very rapidly. Very rapid wetting of the sample occurred, followed by
almost immediate release of MA from all mixtures. The 1:1 (w/w) and
1:2 (w/w) mixtures initially exhibited almost identical release rate while
the 1:1 mixture exhibited a slight slowdown later during the experiment,
but all of the drug substance was released finally in both cases. By
comparing the dissolution tests of the PMs and COMs, it is apparent that
the presence of MG in the mixture greatly accelerated the release of MA,
but this rate was much higher after milling.
Mixtures comprising PVP (Fig. 2) showed slower dissolution than
those based on MG. Probably, a combination of the PVP neutral nature
and good solubility in water caused just little improvement in the case of
MA, whereas there was a greater improvement for IND. In the case of

A) B)
100 100

80 80

60 60
wIND (%)
wIND (%)

40 40
1:4 1:4
1:2 1:2
20 1:1 20 1:1
IND milled IND milled
IND pure IND pure
0 0
0 5 10 15 20 25 30 0 5 10 15 20 25 30

t (min) t (min)
C) D)
100 1:4
100 1:4
1:2 1:2
1:1 1:1
80 MA milled
80 MA milled
MA pure MA pure
60 60
wMA (%)
wMA (%)

40 40

20 20

0 0
0 5 10 15 20 25 30 35 40 45 0 5 10 15 20 25 30 35 40 45

t (min) t (min)

Fig. 2. Dissolution profiles of mixtures of indomethacin (IND) or mefenamic acid (MA) with povidone (PVP) prepared by physical mixing (PM) or co-milling (COM):
IND/PVP PMs (A), IND/PVP COMs (B), MA/PVP PMs (C) and MA/PVP COMs (D).

4
M. Slámová et al.
Fig. 3. XRD diffractograms of mixtures of indomethacin (IND) or mefenamic acid (MA) with meglumine (MG) or povidone (PVP) prepared by physical mixing (PM)
or co-milling (COM) in ratio 1:4: IND/MG (A), MA/MG (B), IND/PVP (C) and MA/PVP (D).
PMs, dissolution rates were very slow. Probably PVP as a large molecule
was likely to coat the IND surface partially, swelled and therefore slowed
drug release into the bulk solution (Fini et al., 2008). However, a notable
improvement was observed in the release rate of IND after co-milling
with PVP as the maximal released fraction plateau was reached within
the first ten minutes of the experiment. This was also shown in a pre­
vious study (Lim et al., 2013a). Interestingly, in the case of MA-PVP
mixtures, the released fraction of MA was higher in PMs as compared
to COMs. In the latter case, the MA released fraction presented only one
quarter of the initial drug load in the flow-through cell, which was not
favorable in terms of short-term dissolution. However, MA would be
probably dissolved within a few hours as the time profiles of the released
fractions were still increasing.
The relative release rate r reached its maximum value (rMAX)
immediately or after some time, and thereafter it decreased with time as
the drug was released in the flow-through cell. The maximum values of
release rates are shown in Table 1. The milling of IND enhanced the rMAX
three times more in comparison to the rMAX of the non-milled drug form.
The mechanically treated MA did not show any significant improvement
in release rates in comparison to the non-milled form. The rMAX reached
a similar, very little values in both drug forms. Co-milling of IND with
MG resulted in clear improvements in the rMAX as seen in Table 1. The
IND was released faster from COMs and PMs, compare to the self-milled
IND. The MG fraction did not have a large effect on rMAX since maximum
r reached similar values for all mixtures (within observed standard de­
viations). All means except that of PM 1:1 were shown to be not
significantly different. Considerable improvement in the dissolution
behavior of MA was also seen. For all mixtures of MA-MG, rMAX values
were much higher compared to the pure or milled MA. Not so positive
5
International Journal of Pharmaceutics 597 (2021) 120312
results were visible for mixtures comprising PVP, as rMAX reached very
similar values for all samples. In the case of MA, co-milling was even less
effective than the PMs. Co-milling of IND with PVP only slightly
increased the rMAX according to the results in Table 1. Moreover, the
amount of PVP in the mixture had a visible influence on the maximum
release rate, which increased with the higher percentage of PVP.
3.2. XRD (crystallinity)
XRD was used for evaluation of crystallinity in the mixtures and also
to verify whether the same crystalline form was obtained after the me­
chanical treatment or not. This was particularly an aspect in the case of
MA with MG, where the acidic drug and small-molecular basic additive
could theoretically have resulted in a salt/co-crystal or a co-amorphous
form. XRD diffractograms of the mixtures with MG are depicted in
Fig. 3A (IND-MG) and 3B (MA-MG). Diffractograms with PVP are
depicted in Fig. 3C (IND-PVP) and 3D (MA-PVP). The results are
depicted for mixtures containing 80% of co-former due to the biggest
influence of co-former on prepared mixtures (more spectra are shown in
the supplementary document). Similar crystallinity in the sample was
found for the physical and co-milled mixture with visible signs of partial
amorphization after milling. The diffractogram of acidic drug and basic
MG, confirmed that no salt was formed (neither for indomethacin nor in
the case of mefenamic acid).
Polymorphic forms of components were clarified by comparing with
the Cambridge Structural Database Refcodes – Indomethacin (INDMET),
Mefenamic acid (XYANAC) and meglumine (NUYJEW). Throughout the
experiments (during milling and co-milling), the polymorphic forms
remained the same and were not converted to another polymorph.
M. Slámová et al. International Journal of Pharmaceutics 597 (2021) 120312

identical, and therefore the ball milling process alone seemed to not
A) B) affect the FTIR spectra, and any shifts or broadening of the peaks are
MA:MG 1:4 COM COM caused by mixing or co-milling of MA with MG. In both cases (PM and
COM), there was a visible interaction between the components. A weak

Intensity (a.u.)
Intensity (a.u.)

PM
MA:MG 1:4 PM
interaction was observed in PM and this interaction was significantly
MG milled MG milled enhanced after co-milling. A new “band” was formed around 1375 cm− 1
(there are visible bands that do not belong neither MA nor MG), indi­
MG pure MG pure
cating chemical interaction between the carbonyl group of MA and the
MA milled MA milled
secondary amino group of MG (Fig. 4B). This interaction may also
MA pure contribute to the improvement of the dissolution profiles of MA with
MA pure
MG.
500 1000 1500 2000 2500 3000 3500 4000 1300 1350 1400 1450

Wavenumber (cm ) -1 Wavenumber (cm-1) 3.4. Particle size distribution

Fig. 4. FTIR spectra for pure and milled mefenamic acid (MA) and meglumine
The morphology of pure substances was monitored by an optical
(MG) and their mixtures (in ratio 1:4) prepared by physical mixing (PM) and co-
microscope. Fig. 5 shows images of pure IND, MA, MG, and PVP. In the
milling (COM).
case of pure IND (Fig. 5A) and pure MA (Fig. 5B), the particles were
irregular, and the particle size was up to approximately 130 µm (IND)
3.3. FTIR
and 60 µm (MA). MG (Fig. 5C) particles were needle shape and up 300
µm (length). PVP particles (Fig. 5D) were approximately spherical and
FTIR studies were conducted to further investigate the nature and
about 50 µm diameter.
extent of interactions between drug and co-former. FTIR study revealed
The differences between PM and COM were monitored by SEM
differences in short-range molecular interactions between MA and co-
(Fig. 6) for mixtures 1:2 (w/w) with MG. The particle size distribution
former in PM and COM. Mixtures of MA with MG, in particular a
(PSD) was measured for mixtures MA-MG 1:2 (Fig. 7). PSD of MG
mixture containing 80% MG, have been chosen for a more in-depth
mixtures (Fig. 7) could not be reliably measured (by laser diffraction)
description since it exhibited the most interesting release changes.
since agglomerates were already formed during the measurement, and
More data showed low or absent effect and some of them are therefore
thus the measured values were not very conclusive. However, as there
shown in the supplementary document. In Fig. 4A the spectrum of pure
was no increase in the initial dissolution rate, the effect of separated
MA shows a signal N-H stretching vibration of secondary amine group at
milled particles was not likely. It has been confirmed from the SEM
3310 cm− 1, C-H stretching of the aliphatic CH3 group at 1445 cm− 1, the
(Fig. 6) that after co-milling, no such milled particles were visible, but
dual signal of C = O stretching at 1645 cm− 1 and C = C stretching of the
rather agglomerates were formed which can significantly affect the
aromatic ring at 1575 cm− 1. This is in line with previous finding in the
dissolution of the drug. This aggregation process can occur spontane­
literature (Kang et al., 2015; Panchagnula et al., 2004). The meglumine
ously in powders because of adhesion forces that always appeared
spectrum also shows the dual signal N-H stretching vibration of sec­
among fine particles obtained by co-milling (Li et al., 2017). Therefore,
ondary amine group at 3237 and 3317 cm− 1, vibration of OH groups at
the prepared mixtures were not primarily monitored for particle size,
2674–2984 cm− 1, C-H stretching of CH2 and CH3 groups at 1431 cm− 1.
but rather for particle structure, interfacial forces and other properties
The spectra of pure MG and MA and after ball milling were almost
associated with the formation of agglomerates. The formation of

Fig. 5. Images from optical microscope for pure substances (scale 50 µm): indomethacin (A), mefenamic acid (B), meglumine (C), povidone (D).

6
M. Slámová et al. International Journal of Pharmaceutics 597 (2021) 120312

Fig. 6. SEM images for mixtures of indomethacin (IND) or mefenamic acid (MA) with meglumine (MG) in ratio 1:2 prepared by physical mixing (PM) or co-milling
(COM): MA/MG PM (A), MA/MG COM (B), IND/MG PM(C), IND/MG COM (D).

agglomerates results in comparatively long-term surface activation, dissolution in several ways - particle size reduction, surface roughness
which helps to accelerate in situ the release of drug (while there is no (process induced disorder), and surface activation. The particle size
increase in the initial dissolution rate) (Loh et al., 2014). determines the comparison of the PM and COM curves but does not
explain the effect of co-milling. The rugged particles, when dissolving,
slowly produced coated, smooth particles. The µ decreased due to the
3.5. Specific release rate
decrease in the specific surface area of the particles. Surface activation
by milling produces composites in which the µ curve decreased only very
The specific release rate (µ) represents the surface equivalent resid­
slowly (Fig. 8A, B). The composite was better formed if the drug tended
ual mass of the non-dissolved drug (Slámová et al., 2020). In an ideal
to form a partially amorphous phase or surface disorder (Fig. 8A). Much
case, the profile of µ would be constant during the entire dissolution.
was evident from the mixtures containing MG (Fig. 8), in the case of
However, this simple assumption is valid only for homogenous material
mixtures with PVP (Fig. 9), the characteristic shape of the µ curves was
composed of mono-sized spherical particles. In real observations, this
also visible, but other influences also played a role and such a system
parameter usually increases faster in the beginning due to the wetting of
was more complicated.
the powder material introduced into the dissolution cell. After reaching
The µ values of IND mixtures with MG showed (Fig. 8A,C) that a
the limit value of released drug fraction representing the maximal µ,
considerable part of the drug was released from the mixture under the
values decrease with the released drug ratio until a minimum is reached
high release rate. Considerable improvement in the dissolution behavior
due to the gradual diminution of a finer fraction of the polydisperse drug
of MA was seen also in µ as depicted in Fig. 8B, D. The MA was released
particles remaining in the system. The drug washout behavior is briefly
from the system with MG a hundred times faster than in the case of the
depicted with the dependency of the µ on the released fraction of drug.
milled drug without the additive. Moreover, almost one half of the initial
Both drugs varied noticeably with the profiles of the µ as seen in Figs. 8
drug amount was released under the high specific release rate in the
and 9.
mixture ratio 1:4 (w/w), and therefore the entire MA was almost
SEM images (Fig. 6) confirm that after co-milling, no separated
released during the first fifteen minutes of the experiment (Fig. 1), which
particles are present in the mixture, but new agglomerates are formed
can be considered as a great enhancement. Mechanistic insights of these
which have long-term surface activation. Milling can accelerate

7
M. Slámová et al.
Fig. 7. Particle size distribution of mefenamic acid (A), meglumine (B) and their mixtures in ratio 1:2 prepared by physical mixing (PM) or co-milling (COM) (C).
positive results were gained from XRD measurements, which showed
that the co-milling MG with MA decreased the crystallinity of the drug
and hence increased the amorphous content in the system (Fig. 3). FTIR
results (Fig. 4) for mixtures comprising MA and MG showed molecular
interaction between molecules, which contributed to the improvement
of MA dissolution.
4. Conclusion
This study directly compared the effect of co-milling with different
co-formers on drugs having different glass forming ability. Our results
showed that the glass forming ability of a drug has a crucial impact on
the dissolution performance of milled and co-milled formulations
thereof. Glass formers are likely to amorphize even when milled sepa­
rately, while non-glass formers require the co-milling with a suitable co-
former to achieve a pronounced effect. It has also been confirmed that
the proper choice of the excipient is the key to maximizing the disso­
lution rates. Specifically, the meglumine was found to contribute enor­
mously to the rapid release of two model NSAID drugs – indomethacin
and mefenamic acid – the effect being almost tenfold as compared to
that of polyvinylpyrrolidone. The glass forming ability of the drug
further affects the properties of composite particles obtained by co-
milling. While the full amorphization or solid phase change did not
occur in studied systems, the surface activation of drug particle was
achieved, improving the dissolution rate. In the case of the studied glass
former, this surface activation was remarkably a long-term effect so the
composition maintained a high specific dissolution rate until a very high
dissolved fraction was achieved. Conversely, the tendency to form such
an active composite was significantly lower for the non-glass former
drug, and the specific dissolution rate declined during the dissolution
process. This observation may indicate the glass former systems can be
8
International Journal of Pharmaceutics 597 (2021) 120312
effectively enhanced by surface activation only, while non-glass forming
systems would require more substantial whole-volume change for the
effective dissolution enhancement. However, further research studies
will be needed to draw final conclusions on comparing various drugs of
different glass-forming ability in the course of co-milling.
CRediT authorship contribution statement
Michaela Slámová: Investigation, Writing - original draft, Visuali­
zation, Funding acquisition, Data curation. Kateřina Prausová: Inves­
tigation. Julie Epikaridisová: Investigation, Validation. Jana
Brokešová: Investigation, Resources. Martin Kuentz: Conceptualiza­
tion, Methodology, Writing - review & editing. Jan Patera: Supervision,
Writing - review & editing. Petr Zámostný: Conceptualization, Meth­
odology, Supervision, Writing - review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgements
This research was supported by MSMT No. 21-SVV/2019. This study
was supported by the Funding Agency of Charles University under Grant
No. 268120/2020 and the Funding Agency of Charles University under
Grant No. SVV 260 547. The work was supported by The Parc. The
authors would like to acknowledge Ing. Martina Kohoutková, Ph.D. for
cooperation with XRD analysis.
M. Slámová et al. International Journal of Pharmaceutics 597 (2021) 120312

Fig. 8. Specific release rates for mixtures of indomethacin (IND) or mefenamic acid (MA) with meglumine (MG) prepared by physical mixing (PM) or co-milling
(COM): IND/MG COM (A), MA/MG COM (B), IND/MG PM (C), MA/MG PM (D).

Fig. 9. Specific release rates for mixtures of indomethacin (IND) or mefenamic acid (MA) with povidone (PVP) prepared by physical mixing (PM) or co-milling
(COM): IND/PVP COM (A), MA/PVP COM (B), IND/PVP PM (C), MA/PVP PM (D).

9
M. Slámová et al.
Appendix A. Supplementary material
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.ijpharm.2021.120312.
References
Alhalaweh, A., Alzghoul, A., Kaialy, W., Mahlin, D., Bergström, C.A.S., 2014.
Computational predictions of glass-forming ability and crystallization tendency of
drug molecules. Mol. Pharm. 11 (9), 3123–3132. https://doi.org/10.1021/
mp500303a.
Bahl, D., Hudak, J., Bogner, R.H., 2008. Comparison of the ability of various
pharmaceutical silicates to amorphize and enhance dissolution of indomethacin
upon co-grinding. Pharm. Dev. Technol. 13 (3), 255–269. https://doi.org/10.1080/
10837450802012869.
Baird, J.A., Van Eerdenbrugh, B., Taylor, L.S., 2010. A classification system to assess the
crystallization tendency of organic molecules from undercooled melts. J. Pharm. Sci.
99 (9), 3787–3806. https://doi.org/10.1002/jps.22197.
Borba, P.A.A., Pinotti, M., De Campos, C.E.M., Pezzini, B.R., Stulzer, H.K., 2016. Sodium
alginate as a potential carrier in solid dispersion formulations to enhance dissolution
rate and apparent water solubility of BCS II drugs. Carbohydr. Polym. 137, 350–359.
https://doi.org/10.1016/j.carbpol.2015.10.070.
Caron, V., Tajber, L., Corrigan, O.I., Healy, A.M., 2011. A comparison of spray drying and
milling in the production of amorphous dispersions of sulfathiazole/
polyvinylpyrrolidone and sulfadimidine/polyvinylpyrrolidone. Mol. Pharm. 8 (2),
532–542. https://doi.org/10.1021/mp1003674.
Fini, A., Cavallari, C., Ospitali, F., 2008. Raman and thermal analysis of indomethacin/
PVP solid dispersion enteric microparticles. Eur. J. Pharm. Biopharm. 70 (1),
409–420. https://doi.org/10.1016/j.ejpb.2008.03.016.
Gupta, M.K., Vanwert, A., Bogner, R.H., 2003. Formation of physically stable amorphous
drugs by milling with neusilin. J. Pharm. Sci. 92 (3), 536–551. https://doi.org/
10.1002/jps.10308.
Isaac, J., Ganguly, S., Ghosh, A., 2016. Co-milling of telmisartan with poly(vinyl alcohol)
- An alkalinizer free green approach to ensure its bioavailability. Eur. J. Pharm.
Biopharm. 101, 43–52. https://doi.org/10.1016/j.ejpb.2016.01.016.
Kaminska, E., Adrjanowicz, K., Kaminski, K., Wlodarczyk, P., Hawelek, L.,
Kolodziejczyk, K., Tarnacka, M., Zakowiecki, D., Kaczmarczyk-Sedlak, I., Pilch, J.,
Paluch, M., 2013. A new way of stabilization of furosemide upon cryogenic grinding
by using acylated saccharides matrices. The role of hydrogen bonds in
decomposition mechanism. Mol. Pharm. 10 (5), 1824–1835. https://doi.org/
10.1021/mp300606p.
Kang, N., Lee, J., Choi, J.N., Mao, C., Lee, E.H., 2015. Cryomilling-induced solid
dispersion of poor glass forming/poorly water-soluble mefenamic acid with
polyvinylpyrrolidone K12. Drug Dev. Ind. Pharm. 41 (6), 978–988. https://doi.org/
10.3109/03639045.2014.920024.
Kawabata, Y., Wada, K., Nakatani, M., Yamada, S., Onoue, S., 2011. Formulation design
for poorly water-soluble drugs based on biopharmaceutics classification system:
Basic approaches and practical applications. Int. J. Pharm. 420 (1), 1–10. https://
doi.org/10.1016/j.ijpharm.2011.08.032.
Koh, I.-B., Shin, S.-C., Lee, Y.-B., 1986. Enhanced dissolution rates of piroxicam from the
ground mixtures with chitin or chitosan. Arch. Pharm. Res. 9 (1), 55–61. https://doi.
org/10.1007/BF02857708.
Kürti, L., Kukovecz, Á., Kozma, G., Ambrus, R., Deli, M.A., Szabó-Révész, P., 2011. Study
of the parameters influencing the co-grinding process for the production of
10
International Journal of Pharmaceutics 597 (2021) 120312
meloxicam nanoparticles. Powder Technol. 212 (1), 210–217. https://doi.org/
10.1016/j.powtec.2011.05.018.
Li, J., Yang, Y., Zhao, M., Xu, H., Ma, J., Wang, S., 2017. Improved oral bioavailability of
probucol by dry media-milling. Mater. Sci. Eng. C 78, 780–786. https://doi.org/
10.1016/j.msec.2017.04.141.
Lim, R.T.Y., Ng, W.K., Tan, R.B.H., 2013a. Dissolution enhancement of indomethacin via
amorphization using co-milling and supercritical co-precipitation processing.
Powder Technol. 240, 79–87. https://doi.org/10.1016/j.powtec.2012.07.004.
Lim, R.T.Y., Ng, W.K., Widjaja, E., Tan, R.B.H., 2013b. Comparison of the physical
stability and physicochemical properties of amorphous indomethacin prepared by
co-milling and supercritical anti-solvent co-precipitation. J. Supercrit. Fluids 79,
186–201. https://doi.org/10.1016/j.supflu.2013.02.017.
Löbmann, K., Flouda, K., Qiu, D., Tsolakou, T., Wang, W., Rades, T., 2014. The influence
of pressure on the intrinsic dissolution rate of amorphous indomethacin.
Pharmaceutics 6, 481–493. https://doi.org/10.3390/pharmaceutics6030481.
Loh, Z.H., Samanta, A.K., Sia Heng, P.W., 2014. Overview of milling techniques for
improving the solubility of poorly water-soluble drugs. Asian J. Pharm. Sci. 10 (4),
255–274. https://doi.org/10.1016/j.ajps.2014.12.006.
Maggi, L., Bruni, G., Maietta, M., Canobbio, A., Cardini, A., Conte, U., 2013. II.
Technological approaches to improve the dissolution behavior of nateglinide, a
lipophilic insoluble drug: Co-milling. Int. J. Pharm. 454 (1), 568–572. https://doi.
org/10.1016/j.ijpharm.2013.06.085.
Maggi, L., Canobbio, A., Bruni, G., Musitelli, G., Conte, U., 2015. Improvement of the
dissolution behavior of gliclazide, a slightly soluble drug, using solid dispersions.
J. Drug Deliv. Sci. Technol. 26, 17–23. https://doi.org/10.1016/j.jddst.2015.01.002.
Mura, P., Cirri, M., Faucci, M.T., Ginès-Dorado, J.M., Bettinetti, G.P., 2002. Investigation
of the effects of grinding and co-grinding on physicochemical properties of
glisentide. J. Pharm. Biomed. Anal. 30 (2), 227–237. https://doi.org/10.1016/
S0731-7085(02)00252-2.
Nkansah, P., Antipas, A., Lu, Y., Varma, M., Rotter, C., Rago, B., El-Kattan, A., Taylor, G.,
Rubio, M., Litchfield, J., 2013. Development and evaluation of novel solid
nanodispersion system for oral delivery of poorly water-soluble drugs. J. Control.
Release 169 (1-2), 150–161. https://doi.org/10.1016/j.jconrel.2013.03.032.
Panchagnula, R., Sundaramurthy, P., Pillai, O., Agrawal, S., Raj, Y.A., 2004. Solid-state
characterization of mefenamic acid. J. Pharm. Sci. 93 (4), 1019–1029. https://doi.
org/10.1002/jps.20008.
Patterson, J.E., James, M.B., Forster, A.H., Lancaster, R.W., Butler, J.M., Rades, T., 2007.
Preparation of glass solutions of three poorly water soluble drugs by spray drying,
melt extrusion and ball milling. Int. J. Pharm. 336 (1), 22–34. https://doi.org/
10.1016/j.ijpharm.2006.11.030.
Slámová, M., Školáková, T., Školáková, A., Patera, J., Zámostný, P., 2020. Preparation of
solid dispersions with respect to the dissolution rate of active substance. J. Drug
Deliv. Sci. Technol. 56, 101518. https://doi.org/10.1016/j.jddst.2020.101518.
Szafraniec, J., Antosik, A., Knapik-Kowalczuk, J., Kurek, M., Syrek, K., Chmiel, K.,
Paluch, M., Jachowicz, R., 2017. Planetary ball milling and supercritical fluid
technology as a way to enhance dissolution of bicalutamide. Int. J. Pharm. 533 (2),
470–479. https://doi.org/10.1016/j.ijpharm.2017.03.078.
Varghese, S., Ghoroi, C., 2017. Improving the wetting and dissolution of ibuprofen using
solventless co-milling. Int. J. Pharm. 533 (1), 145–155. https://doi.org/10.1016/j.
ijpharm.2017.09.062.
Wyttenbach, N., Kirchmeyer, W., Alsenz, J., Kuentz, M., 2016. Theoretical considerations
of the prigogine-defay ratio with regard to the glass-forming ability of drugs from
undercooled melts. Mol. Pharm. 13 (1), 241–250. https://doi.org/10.1021/acs.
molpharmaceut.5b0068810.1021/acs.molpharmaceut.5b00688.s001.